Quantification of soluble phase analytes represents one of the most commonly used techniques applied to a broad range of samples in both basic and clinical immunology laboratories, as well as in context of drug development and diagnostic programs. The recent increase in the application of multiplex immunoassays, such as Luminex, has resulted in a growing array of commercially available multiplex kits. Validated, highly sensitive, and precise methods for such quantification is critical, especially when applied to precious sample collections. While vendors are expected to carry out kit performance validation, discrepancies between technical specifications provided with multiplex kits and their actual performance can be relatively common. Here we present a validation strategy that will aid users to select the optimal kits for their purpose and most validly interpret results from the multiplex assays. To illustrate key considerations when validating and comparing kits, we assess the performance of three conventional multiplex cytokine kits. Our findings confirm the importance of validating the performance of commercial multiplex kits and provide a practical and cost-effective approach that can be readily implemented in both academic and translational laboratory settings.